Central insulin receptors will be characterized pharmacologically, kinetically and biochemically. Optimal conditions for the in vitro assay of 123I-insulin binding to rat brain membranes will be established. The properties of the receptors will be determined by both kinetic (association and dissociation rates) and equilibrium (dose response curves, Scatchard plots and Hill plots) analyses. The central insulin receptor will be solubilized using non-ionic detergents and will be isolated using affinity chromatography. The properties of the soluble receptor will be compared to those of the membrane-bound receptor. Similar studies will be carried out using hepatic membranes, and the properties of peripheral and central insulin receptors will be compared. These experiments will address the possibility that there are multiple forms of insulin receptors. The in vitro binding assay will be used to compare the cellular and regional localization and the time course of development of central insulin receptors. The number and affinity of insulin receptors will be measured in regions of the rat brain. Insulin levels will be measured in the same areas by a double antibody radioimmunoassay. If insulin receptors are involved in neurotransmission, it is likely that both insulin and insulin receptors will be distributed assymetrically. If, however, these receptors are localized on nonneuronal elements such as glial cells or capillaries, their distribution is likely to be relatively uniform. The neuronal or nonneuronal localization of central insulin receptors will be determined using lesions and cell fractionation techniques. Kainic acid, electrolytic, and knife-cut lesions will be used to destroy certain brain nuclei and pathways. Rats made hypoinsulinemic by treatment with streptozotocin will be used to examine insulin receptors in brain areas exposed to or protected from circulating insulin. Biochemical fractionation techniques will be employed to obtain neuron, glia and capillary-enriched fractions with which to carry out 123I-insulin binding. The results of these studies should compliment those obtained in studies of neuronal lesions. Putative functions of insulin in the brain will also be evaluated. The possibility that insulin inhibits or stimulates phosphorylation of brain proteins will be determined. For these studies SDS slab gels will be used to isolate the proteins. The effects of insulin on electrophysiological properties in hippocampal and hypothalamic slices will be determined. The ultimate goal of these studies is to understand the regulation and function of insulin and insulin receptors in the mammalian brain.